Inhibitory Effect Of BD0801 Targeting VEGF/VEGFR Pathway On The Proliferation Of HCC Cell Line Bel7402 And Angiogenesis In Vitro

Objective:To investigate the influence of humanized anti-VEGF monoclonal antibody BD0801 or BD0801 combined with 5-fluorouracil(5-FU)/oxaliplatin(OXA) on the inhibition effect of human hepatocellular cell line Bel7402 proliferation and human microvascular endothelial cell line(HMEC-1) cord formation in vitro.Methods:In vitro, with Bevacizumab as posititive agent, cell proliferation in different concentration of Bel7402 and HMEC-1 was analyzed by 3-(4,5-dimethylthiazole-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay. The DAPI and the flow cytometry assay were applied to detect the changes of apoptotic rate and cell-cycle. Analyses of VEGF content in cell media, and VEGF and pVEGFR expressions were obversed by enzyme-linked immunosorbent assay (ELISA) and Western blot, respectively. The wound healing and Transwell assays were used to investigate effect on the migration of HMEC-1. Tube formation assay was done to test influcence on the angiogenesis of HMEC-1.Results:In vitro, BD0801 targeting VEGF/VEGFR pathway could effectively inhibit proliferation of Bel7402 and HMEC-1. Significant apoptosis-inducing effect was found in FCM. the cell cycle was arrested at Go/G1 phase, and the level of VEGF in cell culture medium was down-regulated. Meanwhile. migration(wound healing and transwell assays) and assembly into vascular structures (tube formation assay) were significantly reduced by incubation with BD0801. The synergistic interaction between BD0801 and 5-FU/OXA was more effective than when the agents used alone. The treatment effect, including cell proliferation, apoptosis and cell cycle arrest, on BD0801 combined with 5-FU/OXA groups was futher strengthened. There was a stronger decrease in VEGF release and the protein expresstion of VEGF and pVEGFR. In addtition, the combined medication groups were remarkably inhibit the migration and angiogenesis of HMEC-1. Compared with Bev, BD0801 monotherapy or combined with 5-FU/OXA groups showed stronger effects in above.Conclusions:First, BD801 targeting VEGF/VEGFR could inhibit cell proliferation, induce apoptosis and arrest cell cycle in Bel7402 and HMEC-1, as well as inhibit angiogenesis of HMEC-1 in vitro. Second, BD0801 in combination with 5-FU/OXA showed synergistic effect of inhibiting cell proliferation, inducing apoptsis, arresting cell cycle as well as inhibiting VEGF/VEGFR pathway on the two cell lines and angiogenesis of HEMC-1. Third, BD0801 has stronger inhibitory effect than Bev. It is suggested that BD0801 might be a potential and promising agent of anti-VEGF monoclonal antibody to be further study in HCC therapy.